Protease enzymes mediate many biological processes, e.g., by editing a polypeptide to a shorter, active form, or by terminating biological activity through degradation of an active polypeptide. Other protease enzymes are concerned with tissue remodeling.
Proteases hydrolyze the amide backbone of polypeptides and during this hydrolysis, a tetrahedral intermediate is formed as part of the enzyme substrate complex. Some analogs of the tetrahedral intermediate can inhibit protease enzymes. Elements other than carbon, specifically, phosphorous and boron, have been used to prepare transition state analogs. Phosphorous: Kam, C. -M.; Nishino, N.; Powers, J. C., xe2x80x9cInhibition of Thermolysin and Carboxypeptidase A by Phosphoramidatesxe2x80x9d, Biochemistry 18, 3032-3038 (1979). Boron: Amiri, P.; Lindquist, R. N.; Matteson, D. S.; Sadhu, K. M. xe2x80x9cBenzamidomethaneboronic Acid: Synthesis and Inhibition of Chymmotrypsinxe2x80x9d, Arch. Biochem. Biophys. 234, 531-536 (1984). There has been only one attempt, however, to utilize silanols in transition state analogs because silanediols have a strong proclivity to self condense and form siloxanes or silicones. The simplest silanediol, dimethylsilanediol, was tested as an inhibitor of angiotensin-converting enzyme and found to be inactive. Galardy, R. E.; Kortylewicz, Z. P. xe2x80x9cInhibitors of angiotensin-converting enzyme containing a tetrahedral arsenic atomxe2x80x9d, Biochem. J. 226, 447-454 (1985). In addition, known silanediols are virtually all dialkyl or diaryl homologues. Lickiss, P. D., xe2x80x9cThe Synthesis and Structure of Organosilanolsxe2x80x9d, Adv. Inorg. Chem. 42, 147-262 (1995). Therefore, organic silanols have been absent from the field of protease inhibition.
It is an object of the invention to provide siliconxe2x80x94containing enzyme inhibitors.
It is a further object of the invention to provide silanols and silanediols and their siloxane oligomers as bioactive molecules, particularly as inhibitors of hydrolase enzymes.
It is a still further object of the invention to provide a process for the synthesis of silanol and silanediolxe2x80x94based peptide mimics as well as their siloxane oligomers.
It is yet another object to provide a method for inhibiting proteases using silicon-containing peptide analogs.
The siliconxe2x80x94containing compounds of the invention are represented by formula I, formula II or formula III. 
wherein X is OH;
Y is OH, H, lower alkyl of one to six carbons with said alkyl preferably methyl, or F;
Z and Zxe2x80x2 are independently H, lower alkyl with said alkyl preferably methyl or ethyl, or Q3Si where Q is lower alkyl with said alkyl preferably methyl or ethyl, or Q is aryl of four to ten carbons with said aryl preferably containing phenyl;
n is preferably 3-50, more preferably 3-10, most preferably 3-5;
nxe2x80x2 is preferably 2-50, more preferably 2-10, most preferably 2-5;
A and B are independently
a) alkyl of one to ten carbons or heteroatoms, preferably three to ten carbons or heteroatoms and said alkyl can be further substituted with aryl;
b) aryl of four to ten carbons or heteroatoms and said aryl can be further substituted with inorganic or organic groups as described below;
c) cyclic of three to ten carbons or heteroatoms; 
xe2x80x83in d, e, and f, CH is bonded to silicon;
R1-R11 groups are each independently hydrogen, alkyl of one to ten carbons or heteroatoms, aryl of four to fourteen carbons or heteroatoms, arylalkyl of five to twenty carbons or heteroatoms; substituted carbonyl or unsubstituted carbonyl.
Heteroatoms are nitrogen, oxygen, silicon or sulfur.
R3, R4, R6, R7, R10 and R11 independently can be one or more naturally-occurring amino acids, e.g., alanine, asparagine, aspartic acid, cysteine, glutamic acid, phenylalanine, glycine, histidine, isoleucine, lysine, leucine, methionine, proline, glutamine, arginine, serine, threonine, valine, tryptophan and tyrosine. Derivatives of these amino acids, as are known in the art, can also be used.
At least one of A or B, or both A and B, are d), e), or f).
By xe2x80x9cindependentlyxe2x80x9d is meant that within formulas I-III, all moieties for the variables such as A, B, R1 to R11, Z and Zxe2x80x2 need not be the same for each variable but may be different moieties within the same compound.
It will also be understood that the compounds have a stable configuration, so that, for example, a destabilizing excess of heteroatoms is not present, and sufficient hydrogens are present to form a stable molecule.
The alkyl groups for A or B may be branched or unbranched and are typically methyl, ethyl, n-butyl, n-propyl, iso-propyl, iso-butyl, iso-pentyl, neo-pentyl, 1-pentyl, 2-pentyl, 3-pentyl, cyclopropylmethyl, and the alkyl groups can be substituted, e.g., with aryl, such as 3-phenyl-1-propyl. The aryl groups for A or B are typically phenyl, phenylmethyl, 1-phenylethyl, 2-phenylethyl, but may also be any other aryl group, for example, pyrrolyl, furanyl, thiophenyl, pyridyl, thiazoyl, imidazoyl, oxazoyl, pyrazinoyl, etc., as well as aryl groups with two or more rings, for example, naphthalenyl, quinolinoyl, isoquinolinoyl, benzothiazoyl, benzofuranyl, etc. The aryl group may also be substituted by an inorganic, alkyl or other aryl group. The cyclic groups for A or B are typically cyclobutylmethyl, cyclopentyl, cyclopentylmethyl, cyclohexyl, cyclohexylmethyl or cycloheptyl.
The alkyl groups for R1 to R11 may be branched or unbranched and contain one to ten members including carbon atoms and optional heteroatoms, preferably three to six members including carbon atoms and optional heteroatoms. Some examples of the alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, isobutyl, heptyl, octyl, nonyl and decyl. The alkyl groups may, in whole or in part, be in the form of rings such as cyclopentyl, cyclohexyl, cycloheptyl, cyclohexylmethyl, tetrahydrofuranyl, tetrahydrothiophenyl, tetrahydropyranyl, piperidinyl, pyrrolindinyl, oxazolindinyl, isoxazolidinyl, etc.
Aryl groups for R1-R11 typically include phenyl, but may also be any other aryl group, for example, pyrrolyl, furanyl, thiophenyl, pyridyl, thiazoyl, imidazoyl, oxazoyl, pyrazinoyl, etc., as well as aryl groups with two or more rings, for example, naphthalenyl, quinolinoyl, isoquinolinoyl, benzothiazoyl, benzofuranyl, etc. The aryl group may also be substituted by an inorganic, alkyl or other aryl group.
The arylalkyl groups for R1-R11 may be any combination of the alkyl and aryl groups described above. These groups may be further substituted. Carbonyl groups for R1-R11 can also be substituted, e.g., with alkyl, aryl, or substitute heteroatoms including oxygen, nitrogen and sulfur.
Alkyl, aryl and cyclic groups in all cases (A, B, R, Z and Zxe2x80x2) can contain one or more double or triple bonds; and/or their hydrogens may be substituted for by inorganic groups such as amino, thio, halo, doubly bonded oxygen (carbonyl) or singly bonded oxygen (hydroxy) or may be substituted for by organic groups such as alkyl, alkenyl or aryl as described herein.
The compounds are stable and can be stored for weeks or longer at room temperature without noticeable decomposition in either solid or solution form. In addition, there is no intrinsic toxicity associated with silicon (Friedberg, K. D. and Schiller, E., Handbook on Toxicity of Inorganic Compounds, Eds. Seiler H. G., and Sigel, H.; Marcel Dekker, New York, 1988, pp. 595-617).
A process is also provided for preparing the compounds of formulas I-III. Preparation of the compounds will generally require a protecting group for the silanol or silanediol that will avoid self condensation. The protecting group must be stable and yet readily removed. Synthesis of the protected silanediol involves formation of siliconxe2x80x94carbon bonds using one or more types of reactions such as those which are described below, followed by deprotection to yield a silanol or silanediol through a reaction generally involving hydrolysis.
The compounds of the invention exhibit pharmaceutical activity and are therefore useful as pharmaceuticals. The compounds of formula I mimic the tetrahedral intermediate of polypeptide hydrolysis and can be incorporated into a polypeptide chain or employed alone or in combinations and used in protease enzyme inhibition. The compounds of formulas II and III are used similarly. Accordingly, a method is provided for inhibiting protease enzymes and in the treatment of related diseases.
Advantageously, the compounds of the invention provide a xe2x80x9ccassettexe2x80x9d which can be inserted into a target peptide or analog of that peptide to result in protease inhibition. Because the compounds are isosteres of the general obligatory tetrahedral intermediate of hydrolysis, protease inhibition using the compounds of the invention is not limited in a choice of target protease.
The invention includes biologically active silanediols, exemplified by Structure II below, which are useful in the design of new drugs. The naturally occurring tetrahedral intermediate of protease mediated hydrolysis is shown in Structure I. 
The Structure II mimics of the tetrahedral intermediate, e.g., when incorporated into a polypeptide chain or used alone or in combination, can be used as highly effective inhibitors of protease enzymes, particularly aspartic proteases (e.g., HIV-I protease and renin) and zinc proteases (e.g., thermolysin and carboxypeptidase A). 
P1 and P1xe2x80x2 are defined as groups on the natural substrate of a protease, or analogs of those groups, that flank the cleavage site of the substrate and are assumed to fit xe2x80x9csubsitesxe2x80x9d on the enzyme (generally referred to as S1 and S1xe2x80x2, respectively) that flank the active site of the enzyme. Additional sites on each side can be specified and are numbered consecutively, e.g. P2, P3, P4, . . . and P2xe2x80x2, P3xe2x80x2, P4xe2x80x2, . . . etc. Schechter, I.; Berger, A. xe2x80x9cOn the Size of the Active Site in Proteases. I. Papain,xe2x80x9d Biochem. Biophys. Res. Commun. 1967, 27, 157-162. Schecter, I.; Berger, A. xe2x80x9cOn the Active Site of Proteases. III. Mapping the Active Site of Papain; Specific Peptide Inhibitors of Papain,xe2x80x9d Biochem. Biophys. Res. Commun. 1968, 32, 898-902.
The silicon-containing compounds of the invention are stable in their configuration and in their activity. Silicon, relative to carbon, has the unique advantage of forming only stable tetrahedral gemxe2x80x94diol (silanediol) and not trigonal silanones. Stable carbonxe2x80x94based gemxe2x80x94diol molecules require electron withdrawing groups at the alpha position to destabilized the trigonal carbonyl and are often in equilibrium with the corresponding carbonyls. This factor and the increased acidity of the silanol as compared with the carbinol, indicates that silanol based enzyme inhibitors can hydrogen bond more strongly to an enzyme active site than carbon based-gemxe2x80x94diols. The term xe2x80x9cgemxe2x80x9d means that two identical substituents are on the same carbon or silicon, e.g., both substituents are hydroxyl groups.
Preferred compounds according to the invention include the following sites: 
The remainder of the molecule is chosen to provide a desired or best set of properties. These properties include enzyme fit, enzyme specificity, solubility, metabolic stability, crystallinity, etc.
Non-limiting examples of compounds of the invention include the following: 
Compounds according to formula I in which A and B are represented by a) and e) are compounds 11, 12, 16, 17 and 20. A compound according to formula I in which A and B are represented by b) and e) is compound 19. A compound according to formula I in which A and B are represented by c) and e) is compound 7. Compound 16 with 
is an example of both a) and f).
Compounds according to formula I in which A and B are represented by d) and e) are compounds 1, 2, 3, 4, 5, 6 and 7. A compound according to formula I in which A and B are both represented by d) is compound 9. Compounds according to formula I in which A and B are both represented by e) are 10, 13, 14, 15, 17, 18. Compounds according to formula I in which A and B are represented by e) and f) are 8 and 16. Compounds 11, 12 and 19 also include e).
A compound according to formula II is 61. A compound according to formula III is 60.
Important considerations in the synthesis were the formation of silicon-carbon bonds, protection to avoid or control oligomerization, removal of the protecting group and hydrolysis of the siliconxe2x80x94containing compound to the silanol, silanediol or siloxane final product.
Since silicon-containing compounds such as silanediols have a proclivity to condense and form siloxanes or silicones, it was necessary to devise a synthesis scheme in which self condensation is inhibited during synthesis. This was achieved by protecting the diol site during synthesis. The choice of protecting group was also important because the protecting group must be capable of being removed under conditions compatible with peptide chemistry. Preferred protecting groups are ones which have unsaturation proximal to a carbonxe2x80x94silicon bond, for example, a phenyl which can also have additional electron donating or withdrawing groups. The protecting groups include substituted or unsubstituted phenyl, vinyl (CHxe2x95x90CH2) and allyl (CH2CHxe2x95x90CH2).
It was determined that triflic (trifluoromethanesulfonic) acid can be used for hydrolysis of the silicon-containing compound to the silanol or silanediol final product. Other acids such as sulfuric acid, hydrofluoric acid, hydrochloric acid, and acetic acid optionally in conjunction with boron trifluoride, can be used, or electrophiles other than H+, such as halogens (chlorine Cl2, bromine Br2, iodinemonochloride ICl), or acid chlorides such as acetylchloride, or electrophiles such as mercuric chloride, can also be used.
Synthesis precursors to the compounds of the invention contain groups attached to silicon that are both generally stable and can be transformed into hydroxyl groups (silanols). These groups can be substituted or unsubstituted aryl, substituted or unsubstituted vinyl, substituted or unsubstituted allyl, substituted or unsubstituted benzyl, or a heteroatomxe2x80x94substituted alkyl, alkoxy or amino group. More specifically, in the synthesis precursors, Aryl includes four to ten carbons and can be substituted. Allyl includes three to ten carbons and can be substituted. Benzyl can also be substituted. Alkyl includes two to four carbons. Alkoxy includes one to four carbons. Substitutions may be by organic or inorganic groups. Inorganic substituents include double-bonded oxygen, i.e., carbonyl, or single bonded oxygen, i.e., hydroxy or alkoxy. Additional inorganic substitutents include amino, thio, halo, etc. Organic substituents include alkyl and aryl. The amines can be primary, secondary or tertiary.
The synthesis of silicon-carbon bonds can be accomplished through various reaction types.
i) As non-limiting examples, these reaction types include nucleophilic attack of a carbon nucleophile, such as a Grignard reagent, on a chlorosilane or alkoxysilane. I. Fleming xe2x80x9cOrganic Silicon Chemistryxe2x80x9d, in Comprehensive Organic Chemistry, D. Barton, W. D. Ollis, Eds. (Pergamon, New York, 1979), vol. 3, pp. 541-686.
For example, a nucleophilic carbon can react with a silicon attached to a leaving group:
R123 Sixe2x80x94X1+R13Mxe2x86x92R123Sixe2x80x94R13
X1 is preferably H, halogen, sulfonate or alkoxy
M=a metal (e.g., Li, Mg, Cu)
The R12 groups are preferably alkyl, aryl or alkoxy.
The reactions are run in an inert solvent (e.g., ether, hexane, toluene) and under an inert atmosphere (e.g., nitrogen, argon) at a temperature between xe2x88x92100xc2x0 C. and +150xc2x0 C. Preferably the reagents are used in a 1:1 ratio, but may range from 1:10 to 10:1.
Compounds such as 1, 2 and 19 can be made using this method.
ii) Alternatively, the opposite arrangement of nucleophile and electrophile can be used, such as a nucleophilic attack by alkyldiphenylsilylcuprate on an iodoalkane.
The silicon can be the nucleophile and carbon the electrophile for example,:
R123Sixe2x80x94M+R13X1xe2x86x92R123Sixe2x80x94R13
Conditions and definitions are as defined in i).
Compounds such as 11 and 12 can be made in this way. iii) An additional method for preparing the desired organosilanes is the hydrosilylation reaction, in which a hydrogenxe2x80x94silicon bond is added across a carbonxe2x80x94carbon double bond, often catalyzed by a metal such as platinum or rhodium. I. Ojima, xe2x80x9cHydrosilylationxe2x80x9d, in The Chemistry of Organic Silicon Compounds; S. Patai and Z. Rappoport, Eds.; Wiley: New York, 1989; Vol. 2; pp 1479-1525.
Hydrosilylation adds a silicon and hydrogen across a carbonxe2x80x94carbon double bond, for example: 
Hydrosilylation reactions are run in an inert solvent (e.g., THF, isopropanol, hexane) and at temperatures between xe2x88x92100xc2x0 C. and +150xc2x0 C. Preferably the reagents are used in a 1:1 ratio, but may range from 1:10 to 10:1. The catalyst can be a radical initiator or a metal. In the case of a radical initiator, from 0.01 to 10 equivalents can be used. Examples of these catalysts are benzoyl peroxide, azo-bis-isobutyronitrile, and organoboranes in the presence of oxygen. In the case of metal catalysts, 0.0001 to 10 equivalents may be used. Various metals can be used, generally platinum or rhodium or cobalt.
Compounds such as 11 and 12 can be made in this way.
iv) Nucleophilic addition of amine (primary or secondary) to alkenylsilanes, usually with base catalysis, can be used, for example: 
R14 through R19 are independently chosen groups, preferably H, or optionally substituted alkyl or aryl. The base is preferably an organometallic reagent such as a Grignard reagent or n-butyllithium and is used in a catalytic amount (0.5 to 0.01 equivalents). One equivalent of the amine is preferably used, but can be used in excess. An inert solvent may be used (ether, hexane).
Compounds such as 16 can be made in this way.
v) Nucleophilic displacement of a halogen by an amine nucleophile can be used, for example: 
X1 is preferably halogen or sulfonate. M is preferably H or a metal (e.g., Li, Na, K, Mg). The moiety NR18R19 is preferably N3 (azide) or phthalimide or succinimide, or R18 and R19 can be H, optionally substituted alkyl or optionally substituted aryl. Preferably a polar, inert solvent is used (e.g., alcohol, ether, DMSO, DMF, THF). The temperature is generally between xe2x88x9250xc2x0 C. and +150xc2x0 C. At least one equivalent of NR18R19 is used, but excess may be employed. When azide is used for NR18R19, the result azide product is reduced to an amine using standard conditions, including but not limited to hydrogenation (e.g., hydrogen gas, platinum catalyst), treatment with thiols, or treatment with lithium aluminum hydride.
Compounds such as 1 and 2 can be made in this way.
vi) Hydrosilylation of an enamine derivative can be used, for example: 
R20 and R21 are preferably H, optionally substituted alkyl or optionally substituted aryl. W1 is preferably a substituted carbonyl derivative such that Nxe2x80x94W1 constitutes an amide, carbamate, or urea. The catalyst is preferably a rhodium derivative such as dirhodium tetraacetate. Preferably the silane and the enamine derivative are used in a ratio of 1:1. Preferably between 0.5 and 0.0001 equivalents of the catalyst is used. The temperature of the reaction is between xe2x88x9250xc2x0 C. and +150xc2x0 C. A reference for this chemistry: Murai, T.; Oda, T.; Kimura, F.; Onishi, H.; Kanda, T.; Kato, S. xe2x80x9cRhodium(II) acetate Catalysed Hydrosilylation of Enamides and N-Vinylureas leading to 1-(Trialkylsilyl)alkylamine Derivatives, xe2x80x9cJ. Chem. Soc., Chem. Commun 2143-2144 (1994).
Compounds such as 3 and 8 can be made in this way.
vii) Silylation of an alpha-metallo amine derivative can be used, for example: 
W2 groups are independently chosen and are preferably a metalation directing group (MUG) or an optionally substituted alkyl or an optionally substituted aryl. R22 and one of the W2 groups can form a ring or both of the W2 groups can form a ring. MDGs are preferably substituted carbonyl groups or substituted imine group or a sulfonyl group or a phosphoryl group. P. Beak, W. J. Zajdel, D. B. Reitz, xe2x80x9cMetalation and Electrophilic Substitution of Amine Derivatives Adjacent to Nitrogen: xcex1-Metallo Amine Synthetic Equivalents,xe2x80x9d Chem. Rev. 84, 471-523 (1984). M is a metal, preferably Li, Na, Mg, or Sn. The temperature of the reaction is preferably between xe2x88x92100xc2x0 C. and +50xc2x0 C.
Compounds such as 10, 13 and 18 can be made in this way.
viii) Rearrangement of alpha-metallo N-silyl compounds can be used, for example: 
W2 group is preferably a metalation directing group (MDG, as defined in (vii), above) or an optionally substituted alkyl or an optionally substituted aryl. M is a metal, preferably Li, Na, Mg, or Sn. The temperature of the reaction is preferably between xe2x88x92100xc2x0 C. and +50xc2x0 C.
Compounds such as 16 and 17 can be made in this way.
Deprotection of the silanol or silanediol generally involves hydrolysis. In the case of phenyltrialkyl or diphenyldialkylsilanes, this is accomplished by treatment with acid to break the siliconxe2x80x94phenyl bond, followed by addition of water to generate the silanol or silanediol. Eaborn, E. xe2x80x9cCleavages of Aryl-10 Silicon and Related Bonds by Electrophiles,xe2x80x9d J. Organomet. Chem. 100, 43-57 (1975).
In synthesis methods i) through viii), R12-R23 can be chosen to provide A, B and/or R1-R11 in the final product. With R12, a further reaction sequence results in X and/or Y in the final product. W1 and W2 will generally be removed. All of the precursor compounds in i)-viii) can be made by methods known in the art or the reagents are commercially available. An example of a synthesis scheme is as follows: 
Difluorodiphenylsilane 24 is alkylated sequentially with 1-bromomagnesium-3-butene and 2-lithio-1,3-dithiane to give 25. Deprotonation of the dithiane 25 with n-butyllithium and alkylation of the resulting anion gives 26. Hydrolysis of dithiane 26 with mercury(II) chloride yields a silaketone which is then reduced with lithium aluminum hydride. The resulting alcohol is derivatized with methanesulfonyl chloride to give the methylsulfonate 27. This sulfonate is then displaced with sodium azide to give an alpha-azido silane that is reduced to an alpha-amino silane with lithium aluminum hydride. The amine is condensed with benzoyl chloride to yield amide 28. Oxidative cleavage of the alkene in 28 to a carboxylic acid is performed with potassium permanganate. The acid is then condensed with benzyl amine using diphenylphosphoryl azide as a dehydrating agent. The resulting diamide 29 is treated with trifluoromethanesulfonic acid in trifluoroacetic acid at 0xc2x0 C. for one hour. Addition of water and extraction of the aqueous phase with dichloromethane yields silanediol 1.
The compounds of the invention inhibit protease enzymes including metallo, apartyl and serine proteases.
Four classes of proteases are known and these are categorized by the catalytic functionality at the active site: aspartic proteases, metalloproteases, serine proteases and cysteine proteases. All four classes contain important therapeutic targets for enzyme inhibition.
Non-limiting examples of therapeutic targets are shown in the table below.
As non-limiting examples, compounds 1, 2 and 13 depicted above can be used to inhibit angiontensin-converting enzyme in the treatment of hypertension. Compounds 3, 8, 11, 12, 21 and 22 can be used to inhibit renin in the treatment of hypertension. Compounds 4, 7, 9, 10, 14, 15, 18 and 20 can be used to inhibit HIV protease in the treatment of AIDS. Compounds 16 and 19 can be used to inhibit elastase in the treatment of emphysema and cystic fibrosis. Compound 17 can be used to inhibit thrombin in the treatment of thrombosis. Compound 5 can be used to inhibit stromelysin in the treatment of arthritis. Compound 6 can be used to inhibit collagenase in the treatment of arthritis.
The naturally-occuring polypeptide cleavage mechanisms of action of the four classes of proteases have been studied.
Aspartic and metallo proteases catalyze the addition of water to the amide bond and stabilize the tetrahedral intermediate of hydrolysis by hydrogen bonding to a pair of aspartic acid residues in aspartic proteases or by coordination to a metal (usually zinc) in metallo proteases.
With aspartic proteases, the catalytic mechanism involves the concerted action of two aspartyl carboxy groups, only one of which is protonated. De Voss, J. J. et al., J. Med. Chem. 37, 665-673 (1994). The protonated aspartyl hydrogen bonds to the amide carbonyl of the substrate and the unprotonated aspartyl to a water molecule. Transfer of the hydrogen from the aspartyl to the carbonyl group of the substrate coupled with addition of a water molecule gives a gem-diol transition-state intermediate. One of the two hydrogens of the water is retained and shared by the aspartyl groups. The tetrahedral gem-diol intermediate is then cleaved, again with the help of the two differently protonated aspartyl groups. Inhibitors of aspartic acid proteases such as renin and HIV-1, have included hydroxyethylene, dihydroxyethylene, xcex1-dicarbonyl, hydroxyethylamine, phosphinate, reduced amide and statine-like groups. Vacca, J. P., xe2x80x9cDesign of Tight-Binding Human Immunodeficiency Virus Type 1 Protease Inhibitorsxe2x80x9d, Methods Enzymol. 241, 311-334 (1994). But there has been no suggestion to use silanols to inhibit aspartic proteases.
The silanol-containing compounds of the invention are isosteres of the hydrated amide bonds that aspartic proteases act upon, but, advantageously, the compounds of the invention are not cleavable under enzymatic conditions. The silanols are tetrahedral in structure and are believed to bind to the aspartic protease enzyme by forming hydrogen bonds to the aspartic acid residues that are present in the enzyme active site. Thus these isosteres function as stable, non-hydrolyzable transition-state mimics (analogs) of the enzyme-catalyzed hydrolysis reaction of the substrate amide bond.
In metalloproteases, a metal coordinates and activates the polypeptide amide carbonyl for nucleophilic attack by water. Carboxypeptidase A and thermolysin are two well studied metalloproteases. Matthews, B. W., Acc. Chem, Res. 21, 333 (1988); Christianson, D. W. and Lipscomb, W. N., Acc. Chem. Res. 22, 62 (1989). Both of these contain zinc at the active site, similar to the clinically important angiotensinxe2x80x94converting enzyme (ACE) (Rich, D. H., xe2x80x9cPeptidase Inhibitors,xe2x80x9d in Comprehensive Medicinal Chemistry, C. Hansch et al., Ed., Pergamon, New York, 1990, pp. 391441) and enkephanlinase enzymes. Other metalloproteases are Endothelin Converting Enzyme (ECE), the matrix metalloproteases (collagenase, stromelysin, gelatinase) and neural endopeptidase.
Inhibitors of metalloproteases have included sites incorporating thiols, aldehydes which can hydrate, hydroxamic acid, carboxylalkylamine, ketone (which can hydrate), phosphinic acid, phosphonamide, phosphonate and aminoketone (which can hydrate). For example, an inhibitor of ACE includes a ketone site (Gordon, E. M. et al., xe2x80x9cKetomethyldipeptides II. Effect of Modification of the xcex1-Aminoketone Portion on Inhibition of Angiotensin Converting Enzymexe2x80x9d, Biochem. Biophys. Res. Commun. 124, 148-155 (1984)) which is expected to be hydrated as the gem-diol. There has been no suggestion to utilize silanols in the inhibition of metalloproteases.
Serine and cysteine proteases utilize a two-step process with an initial nucleophilic attack on an amide carbonyl by a serine or cysteine residue, generating a tetrahedral intermediate of hydrolysis which is covalently attached to the enzyme. These mechanisms of hydrolysis are discussed in detail by R. H. Rich, xe2x80x9cPeptidase Inhibitorsxe2x80x9d in Comprehensive Medicinal Chemistry, P.G. Sommes and J. B. Taylor, eds., Pergamon, New York 1990, Vol. 2, pp. 391441, and G. Fischer, xe2x80x9cTrends in Protease Inhibitionxe2x80x9d, National Product Reports, 1988, 465-495. There has been no suggestion to utilize silanols in the inhibition of serine proteases or cysteine proteases.
Serine proteases include, for example, thrombin and elastase. The mode of action of serine proteases involves the amino acid serine whose alcohol acts as a nucleophile. Inhibitors of serine protease include sites incorporating trifluoromethylketone, aldehyde, boronic acid, xcex1-dicarbonyl, fluoromethylene ketone, borinic acid and phosphonate. In addition, alkylating agents can permanently derivative the serine nucleophile at the enzyme active site. Activated carbonyls or other electrophilic centers interact with the nuclephilic serine oxygen forming a covalent, but not necessarily permanently bound, complex. More specifically, the serine protease, xcex1-lytic protease is inhibited by a peptide compound containing phenyl phosphonate ester 
Bone, R. et al., xe2x80x9cCrystal Structures of xcex1-Lytic Protease Complexes with Irreversibly Bound Phosphonate Estersxe2x80x9d, Biochemistry 30, 2263-2272 (1991). As another example, carbon-based 1,1-diols, such as hydrated trifluoromethylketones also inhibit serine protease (Govardhan, C. P. and Abeles, R. H., xe2x80x9cStructure-Activity Studies of Fluoroketone Inhibitors of xcex1-Lytic Protease and Human Leucocyte Elastasexe2x80x9d, Arch. Biochem. Biophys. 280, 137-146 (1990)). Beginning with 
the inhibitor is dehydrated to the ketone which then reacts with the serine alcohol nucleophile. In the invention, in the inhibition of serine protease enzymes, the oxygens on silicon are exchangeable with the serine alcohol nucleophile. Studies of silane stereochemistry provide convincing evidence for the nucleophilic displacement of oxygen substituents on silicon by alcohols (Corriu, R. J. P. et al., xe2x80x9cStereochemistry at Siliconxe2x80x9d, Topics in Stereochemistry 15, 80-103 (1984)).
In the silicon-containing protease inhibitors of the invention, the carbon-silicon bond is strong and non-hydrolyzable, and the silicon is tetrahedral. Hydroxyl groups on silicon are good hydrogen bond acceptors and are also slightly more acidic than carbinols, making them excellent hydrogen bond donors. They will therefore hydrogen bond to aspartic acid groups in aspartic proteases, and will also act as a chelating group for the metal of metalloproteases. In addition, the hydroxyl groups on silicon are exchangeable with water and will therefore exchange with a serine hydroxyl for serine protease inhibition.
The compounds of the invention are particularly effective, for example, in the inhibition of aspartic proteases HIV-1-protease and renin; metalloproteases ACE, collagenase and stromelysin; and serine proteases thrombin and elastin.
The amount of compound used for inhibition can be determined analogously with known inhibitors of enzymes such as renin or other enzymes listed in TABLE 1 above. Accordingly, the compounds can be used in the treatment of the pathologic conditions such as those listed in TABLE 1. The compounds exhibit antiretroviral activity an can be used to treat retroviral disease such as human immunodeficiency syndrome (AIDS) analogously with the known inhibitors described by Fisher, et al. (Fisher, J. F.; Tarpley, W. G.; Thaisrivongs, S. xe2x80x9cHIV Protease Inhibitors,xe2x80x9d in Design of Enzyme Inhibitors as Drugs; M. Sandler and H.J. Smith, Ed.; Oxford University: New York, 1994; Vol. 2; pp 226-289).
For the pharmaceutical purposes described above, the compounds of the invention can be formulated per se in pharmaceutical preparations or formulated in the form of pharmaceutically acceptable salts, optionally with known pharmaceutically acceptable adjuvants or carriers. These preparations can be prepared according to conventional chemical methods and can be administered enterally, e.g., orally as tablets; parentally, e.g., intravenously intramuscularly or subcutaneously, as injectable solutions or suspensions; or in the form of a spray inhalation.
Pharmaceutical preparations contain a protease-inhibiting effective amount of the compound of formula I, II and/or III. The dosage, analogously with known enzyme inhibiting peptides, depends on the species, body weight, age and mode of administration. The daily dosage is preferably about 0.02-500 mg/kg of body weight per day, more preferably, about 1-20 mg/kg.